Pyrazole compounds, compositions and methods for treatment of degenerative diseases and disorders

ABSTRACT

Provided herein are compounds of the formula (I): 
                         
as well as pharmaceutically acceptable salts thereof, wherein the substituents are as those disclosed in the specification. These compounds, and the pharmaceutical compositions containing them, are useful for the treatment of degenerative diseases and disorders.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is related to U.S. application Ser. No. 13/798,421,filed Mar. 13, 2013; Ser. No. 13/798,383, filed Mar. 13, 2013; and U.S.application Ser. No. 13/636,754, filed Feb. 7, 2013, which claimspriority to PCT/US2011/029846, filed Mar. 24, 2011.

FIELD OF THE INVENTION

The invention is directed to compounds of formula (I):

and to pharmaceutical compositions comprising the compounds. Thecompounds and compositions disclosed herein protect against calcium- andoxidative-stress mediated damage to mitochondrial function and areuseful for the treatment of degenerative diseases and disorders.

All documents cited or relied upon below are expressly incorporatedherein by reference.

BACKGROUND OF THE INVENTION

Mitochondria are cellular organelles present in most eukaryotic cells.One of their primary functions is oxidative phosphorylation, a processthrough which energy derived from metabolism of fuels like glucose orfatty acids is converted to ATP, which is then used to drive variousenergy-requiring biosynthetic reactions and other metabolic activities.Mitochondria have their own genomes, separate from nuclear DNA,comprising rings of DNA with about 16,000 base pairs in human cells.Each mitochondrion may have multiple copies of its genome, andindividual cells may have hundreds of mitochondria. In addition tosupplying cellular energy, mitochondria are involved in a range of otherprocesses, such as signaling, cellular differentiation, cell death, aswell as the control of the cell cycle and cell growth (McBride et al.,Curr. Biol., 2006, 16 (14): R551).

As mitochondria produce ATP, they simultaneously yield reactive oxygenspecies (ROS), which are harmful free radicals that circulate throughoutthe cell, the mitochondria, and the body, causing more damage. Thecirculation of ROS leads to the activation of reactive nitrogencompounds, which in turn induce, or activate, genes in the DNA that areassociated with many degenerative diseases. The DNA for eachmitochondrion (mtDNA) remains unprotected within the membrane of themitochondrion itself. In comparison to the DNA in the nucleus of thecell (nDNA), mtDNA is easily damaged by free radicals and the ROS thatit produces. Freely floating mtDNA lacks protective measures associatedwith nDNA, and therefore suffers from multiple mutations. It has beenestimated that the lack of protective measures results in mutations tomtDNA occurring 10 to 20 times more frequently than mutations to nDNA.

Mitochondrial damage and/or dysfunction contribute to various diseasestates. Some diseases are due to mutations or deletions in themitochondrial genome. Mitochondria divide and proliferate with a fasterturnover rate than their host cells, and their replication is undercontrol of the nuclear genome. If a threshold proportion of mitochondriain a cell is defective, and if a threshold proportion of such cellswithin a tissue have defective mitochondria, symptoms of tissue or organdysfunction can result. Practically any tissue can be affected, and alarge variety of symptoms can be present, depending on the extent towhich different tissues are involved.

A fertilized ovum might contain both normal and genetically defectivemitochondria. The segregation of defective mitochondria into differenttissues during division of this ovum is a stochastic process, as will bethe ratio of defective to normal mitochondria within a given tissue orcell (although there can be positive or negative selection for defectivemitochondrial genomes during mitochondrial turnover within cells). Thus,a variety of different pathologic phenotypes can emerge out of aparticular point mutation in mitochondrial DNA. Conversely, similarphenotypes can emerge from mutations or deletions affecting differentgenes within mitochondrial DNA. Clinical symptoms in congenitalmitochondrial diseases often manifest in postmitotic tissues with highenergy demands like brain, muscle, optic nerve, and myocardium, butother tissues including endocrine glands, liver, gastrointestinal tract,kidney, and hematopoietic tissue are also involved, again depending inpart on the segregation of mitochondria during development, and on thedynamics of mitochondrial turnover over time.

In addition to congenital disorders involving inherited defectivemitochondria, acquired mitochondrial damage and/or dysfunctioncontribute to diseases, particularly neurodegenerative disordersassociated with aging like Parkinson's, Alzheimer's, Huntington'sDiseases. The incidence of somatic mutations in mitochondrial DNA risesexponentially with age; and diminished respiratory chain activity isfound universally in aging people. Mitochondrial dysfunction is alsoimplicated in excitotoxic neuronal injury, such as that associated withseizures or ischemia.

Other pathologies with etiology involving mitochondrial damage and/ordysfunction include schizophrenia, bipolar disorder, dementia, epilepsy,stroke, cardiovascular disease, retinal degenerative disease (e.g.,age-related macular degeneration, Stargardt's disease, glaucoma,retinitis pigmentosa, and optic nerve degeneration), and diabetesmellitus. A common thread thought to link these seemingly-unrelatedconditions is cellular damage causing oxidative stress. Oxidative stressis caused by an imbalance between the production of reactive oxygen anda biological system's ability to readily detoxify the reactiveintermediates or easily repair the resulting damage. All forms of lifemaintain a reducing environment within their cells. This reducingenvironment is preserved by enzymes that maintain the reduced statethrough a constant input of metabolic energy. Disturbances in thisnormal redox state can cause toxic effects through the production ofperoxides and free radicals that damage all components of the cell,including proteins, lipids, and DNA.

Mitochondrial damage and/or dysfunction particularly contribute todegenerative disesaes. Degenerative diseases are diseases in which thefunction or structure of the affected tissues or organs willprogressively deteriorate over time. Some examples of degenerativediseases are retinal degenerative disease, e.g., age-related maculardegeneration, Stargardt's disease, glaucoma, retinitis pigmentosa, andoptic nerve degeneration; amyotrophic lateral sclerosis (ALS), e.g., LouGehrig's Disease; Alzheimer's disease; Parkinson's Disease; multiplesystem atrophy; Niemann Pick disease; atherosclerosis; progressivesupranuclear palsy; cancer; Tay-Sachs disease; diabetes; heart disease;keratoconus; inflammatory bowel disease (IBD); prostatitis;osteoarthritis; osteoporosis; rheumatoid arthritis; and Huntington'sdisease.

Treatment of degenerative diseases involving mitochondrial damage and/ordysfunction has heretofore involved administration of vitamins andcofactors used by particular elements of the mitochondrial respiratorychain. Coenzyme Q (ubiquinone), nicotinamide, riboflavin, carnitine,biotin, and lipoic acid are used in patients with occasional benefit,especially in disorders directly stemming from primary deficiencies ofone of these cofactors. However, while useful in isolated cases, no suchmetabolic cofactors or vitamins have been shown to have general utilityin clinical practice in treating degenerative diseases involvingmitochondrial damage and/or dysfunction.

Therefore, a need exists for new drug therapies for the treatment ofsubjects suffering from or susceptible to the above disorders orconditions associated with mitochondrial damage and/or dysfunction. Inparticular, a need exists for new drugs having one or more improvedproperties (such as safety profile, efficacy or physical properties)relative to those currently available.

SUMMARY OF THE INVENTION

The present invention is directed to compounds of formula I:

wherein:R₁ is lower alkyl, trimethylsilyl or pyridinyl;one of R₂ or R_(2′) is absent and the other is —CH₂R₃ or —CH₂C(O)R₃; andR₃ is pyridinyl, 1H-indol-3-yl, unsubstituted phenyl or phenyl mono-,bi- or tri-substituted independently with alkoxy,or a pharmaceutically acceptable salt thereof.

The present invention is also directed to pharmaceutical compositionscontaining the above compounds, method of using the compounds and tomethods of treating degenerative diseases and disorders.

DETAILED DESCRIPTION OF THE INVENTION

It is to be understood that the terminology employed herein is for thepurpose of describing particular embodiments, and is not intended to belimiting. Further, although any methods, devices and materials similaror equivalent to those described herein can be used in the practice ortesting of the invention, the preferred methods, devices and materialsare now described.

As used herein, the term “alkyl”, alone or in combination with othergroups, refers to a branched or straight-chain monovalent saturatedaliphatic hydrocarbon radical of one to twenty carbon atoms, preferablyone to sixteen carbon atoms, more preferably one to ten carbon atoms.

As used herein, the term “alkenyl”, alone or in combination with othergroups, refers to a straight-chain or branched hydrocarbon residuehaving an olefinic bond.

The term “cycloalkyl” refers to a monovalent mono- or polycarbocyclicradical of three to ten, preferably three to six carbon atoms. This termis further exemplified by radicals such as cyclopropyl, cyclobutyl,cyclopentyl, cyclohexyl, cycloheptyl, norbornyl, adamantyl, indanyl andthe like. In a preferred embodiment, the “cycloalkyl” moieties canoptionally be substituted with one, two, three or four substituents.Each substituent can independently be, alkyl, alkoxy, halogen, amino,hydroxyl or oxygen unless otherwise specifically indicated. Examples ofcycloalkyl moieties include, but are not limited to, optionallysubstituted cyclopropyl, optionally substituted cyclobutyl, optionallysubstituted cyclopentyl, optionally substituted cyclopentenyl,optionally substituted cyclohexyl, optionally substituted cyclohexylene,optionally substituted cycloheptyl, and the like or those which arespecifically exemplified herein.

The term “heterocycloalkyl” denotes a mono- or polycyclic alkyl ring,wherein one, two or three of the carbon ring atoms is replaced by aheteroatom such as N, O or S. Examples of heterocycloalkyl groupsinclude, but are not limited to, morpholinyl, thiomorpholinyl,piperazinyl, piperidinyl, pyrrolidinyl, tetrahydropyranyl,tetrahydrofuranyl, 1,3-dioxanyl and the like. The heterocycloalkylgroups may be unsubstituted or substituted and attachment may be throughtheir carbon frame or through their heteroatom(s) where appropriate.

The term “lower alkyl”, alone or in combination with other groups,refers to a branched or straight-chain alkyl radical of one to ninecarbon atoms, preferably one to six carbon atoms, more preferably one tofour carbon atoms. This term is further exemplified by radicals such asmethyl, ethyl, n-propyl, isopropyl, n-butyl, s-butyl, isobutyl, t-butyl,n-pentyl, 3-methylbutyl, n-hexyl, 2-ethylbutyl and the like.

The term “aryl” refers to an aromatic mono- or polycarbocyclic radicalof 6 to 12 carbon atoms having at least one aromatic ring. Examples ofsuch groups include, but are not limited to, phenyl, naphthyl,1,2,3,4-tetrahydronaphthalene, 1,2-dihydronaphthalene, indanyl,1H-indenyl and the like.

The alkyl, lower alkyl and aryl groups may be substituted orunsubstituted. When substituted, there will generally be, for example, 1to 4 substituents present. These substituents may optionally form a ringwith the alkyl, lower alkyl or aryl group with which they are connected.Substituents may include, for example: carbon-containing groups such asalkyl, aryl, arylalkyl (e.g. substituted and unsubstituted phenyl,substituted and unsubstituted benzyl); halogen atoms andhalogen-containing groups such as haloalkyl (e.g. trifluoromethyl);oxygen-containing groups such as alcohols (e.g. hydroxyl, hydroxyalkyl,aryl(hydroxyl)alkyl), ethers (e.g. alkoxy, aryloxy, alkoxyalkyl,aryloxyalkyl, more preferably, for example, methoxy and ethoxy),aldehydes (e.g. carboxaldehyde), ketones (e.g. alkylcarbonyl,alkylcarbonylalkyl, arylcarbonyl, arylalkylcarbonyl, arycarbonylalkyl),acids (e.g. carboxy, carboxyalkyl), acid derivatives such as esters(e.g. alkoxycarbonyl, alkoxycarbonylalkyl, alkylcarbonyloxy,alkylcarbonyloxyalkyl), amides (e.g. aminocarbonyl, mono- ordi-alkylaminocarbonyl, aminocarbonylalkyl, mono- ordi-alkylaminocarbonylalkyl, arylaminocarbonyl), carbamates (e.g.alkoxycarbonylamino, aryloxycarbonylamino, aminocarbonyloxy, mono- ordi-alkylaminocarbonyloxy, arylminocarbonloxy) and ureas (e.g. mono- ordi-alkylaminocarbonylamino or arylaminocarbonylamino);nitrogen-containing groups such as amines (e.g. amino, mono- ordi-alkylamino, aminoalkyl, mono- or di-alkylaminoalkyl), azides,nitriles (e.g. cyano, cyanoalkyl), nitro; sulfur-containing groups suchas thiols, thioethers, sulfoxides and sulfones (e.g. alkylthio,alkylsulfinyl, alkylsulfonyl, alkylthioalkyl, alkylsulfinylalkyl,alkylsulfonylalkyl, arylthio, arysulfinyl, arysulfonyl, arythioalkyl,arylsulfinylalkyl, arylsulfonylalkyl); and heterocyclic groupscontaining one or more heteroatoms, (e.g. thienyl, furanyl, pyrrolyl,imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, oxadiazolyl,thiadiazolyl, aziridinyl, azetidinyl, pyrrolidinyl, pyrrolinyl,imidazolidinyl, imidazolinyl, pyrazolidinyl, tetrahydrofuranyl, pyranyl,pyronyl, pyridyl, pyrazinyl, pyridazinyl, piperidyl, hexahydroazepinyl,piperazinyl, morpholinyl, thianaphthyl, benzofuranyl, isobenzofuranyl,indolyl, oxyindolyl, isoindolyl, indazolyl, indolinyl, 7-azaindolyl,benzopyranyl, coumarinyl, isocoumarinyl, quinolinyl, isoquinolinyl,naphthridinyl, cinnolinyl, quinazolinyl, pyridopyridyl, benzoxazinyl,quinoxalinyl, chromenyl, chromanyl, isochromanyl, phthalazinyl andcarbolinyl).

The term “heteroaryl,” refers to an aromatic mono- or polycyclic radicalof 5 to 12 atoms having at least one aromatic ring containing one, two,or three ring heteroatoms selected from N, O, and S, with the remainingring atoms being C. One or two ring carbon atoms of the heteroaryl groupmay be replaced with a carbonyl group.

The heteroaryl group described above may be substituted independentlywith one, two, or three substituents. Substituents may include, forexample: carbon-containing groups such as alkyl, aryl, arylalkyl (e.g.substituted and unsubstituted phenyl, substituted and unsubstitutedbenzyl); halogen atoms and halogen-containing groups such as haloalkyl(e.g. trifluoromethyl); oxygen-containing groups such as alcohols (e.g.hydroxyl, hydroxyalkyl, aryl(hydroxyl)alkyl), ethers (e.g. alkoxy,aryloxy, alkoxyalkyl, aryloxyalkyl), aldehydes (e.g. carboxaldehyde),ketones (e.g. alkylcarbonyl, alkylcarbonylalkyl, arylcarbonyl,arylalkylcarbonyl, arycarbonylalkyl), acids (e.g. carboxy,carboxyalkyl), acid derivatives such as esters (e.g. alkoxycarbonyl,alkoxycarbonylalkyl, alkylcarbonyloxy, alkylcarbonyloxyalkyl), amides(e.g. aminocarbonyl, mono- or di-alkylaminocarbonyl, aminocarbonylalkyl,mono- or di-alkylaminocarbonylalkyl, arylaminocarbonyl), carbamates(e.g. alkoxycarbonylamino, aryloxycarbonylamino, aminocarbonyloxy, mono-or di-alkylaminocarbonyloxy, arylminocarbonloxy) and ureas (e.g. mono-or di-alkylaminocarbonylamino or arylaminocarbonylamino);nitrogen-containing groups such as amines (e.g. amino, mono- ordi-alkylamino, aminoalkyl, mono- or di-alkylaminoalkyl), azides,nitriles (e.g. cyano, cyanoalkyl), nitro; sulfur-containing groups suchas thiols, thioethers, sulfoxides and sulfones (e.g. alkylthio,alkylsulfinyl, alkylsulfonyl, alkylthioalkyl, alkylsulfinylalkyl,alkylsulfonylalkyl, arylthio, arysulfinyl, arysulfonyl, arythioalkyl,arylsulfinylalkyl, arylsulfonylalkyl); and heterocyclic groupscontaining one or more heteroatoms, (e.g. thienyl, furanyl, pyrrolyl,imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, oxadiazolyl,thiadiazolyl, aziridinyl, azetidinyl, pyrrolidinyl, pyrrolinyl,imidazolidinyl, imidazolinyl, pyrazolidinyl, tetrahydrofuranyl, pyranyl,pyronyl, pyridyl, pyrazinyl, pyridazinyl, piperidyl, hexahydroazepinyl,piperazinyl, morpholinyl, thianaphthyl, benzofuranyl, isobenzofuranyl,indolyl, oxyindolyl, isoindolyl, indazolyl, indolinyl, 7-azaindolyl,benzopyranyl, coumarinyl, isocoumarinyl, quinolinyl, isoquinolinyl,naphthridinyl, cinnolinyl, quinazolinyl, pyridopyridyl, benzoxazinyl,quinoxalinyl, chromenyl, chromanyl, isochromanyl, phthalazinyl,benzothiazoyl and carbolinyl).

As used herein, the term “alkoxy” means alkyl-O—; and “alkoyl” meansalkyl-CO—. Alkoxy substituent groups or alkoxy-containing substituentgroups may be substituted by, for example, one or more alkyl groups.

As used herein, the term “halogen” means a fluorine, chlorine, bromineor iodine radical, preferably a fluorine, chlorine or bromine radical,and more preferably a bromine or chlorine radical.

Compounds of formula I can have one or more asymmetric carbon atoms andcan exist in the form of optically pure enantiomers, mixtures ofenantiomers such as, for example, racemates, optically purediastereoisomers, mixtures of diastereoisomers, diastereoisomericracemates or mixtures of diastereoisomeric racemates. The opticallyactive forms can be obtained for example by resolution of the racemates,by asymmetric synthesis or asymmetric chromatography (chromatographywith a chiral adsorbents or eluant). The invention embraces all of theseforms.

As used herein, the term “pharmaceutically acceptable salt” means anypharmaceutically acceptable salt of the compound of formula (I). Saltsmay be prepared from pharmaceutically acceptable non-toxic acids andbases including inorganic and organic acids and bases. Such acidsinclude, for example, acetic, benzenesulfonic, benzoic, camphorsulfonic,citric, ethenesulfonic, dichloroacetic, formic, fumaric, gluconic,glutamic, hippuric, hydrobromic, hydrochloric, isethionic, lactic,maleic, malic, mandelic, methanesulfonic, mucic, nitric, oxalic, pamoic,pantothenic, phosphoric, succinic, sulfuric, tartaric, oxalic,p-toluenesulfonic and the like. Particularly preferred are fumaric,hydrochloric, hydrobromic, phosphoric, succinic, sulfuric andmethanesulfonic acids. Acceptable base salts include alkali metal (e.g.sodium, potassium), alkaline earth metal (e.g. calcium, magnesium) andaluminum salts.

In one embodiment of the invention, provided is a compound of formula(I):

wherein:R₁ is lower alkyl, trimethylsilyl or pyridinyl;one of R₂ or R_(2′) is absent and the other is —CH₂R₃ or —CH₂C(O)R₃; andR₃ is pyridinyl, 1H-indol-3-yl, unsubstituted phenyl or phenyl mono-,bi- or tri-substituted independently with alkoxy,or a pharmaceutically acceptable salt thereof.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₁ is lower alkyl.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₁ is trimethylsilyl.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₁ is pyridinyl.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein one of R₂ or R_(2′) is absent and the other is—CH₂R₃.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein one of R₂ or R_(2′) is absent and the other is—CH₂C(O)R₃.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₃ is pyridinyl.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₃ is 1H-indol-3-yl.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₃ is phenyl mono-substituted with methoxy.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₃ is phenyl bi-substituted with methoxy.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₃ is phenyl tri-substituted with methoxy.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein R₂ is absent, R_(2′) is —CH₂C(O)R₃ and R₃ isphenyl bi-substituted with methoxy.

In another embodiment of the invention, provided is a compound accordingto formula (I), wherein the compound is:

-   Ethyl    1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate;-   Ethyl    1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-5-(pyridin-2-yl)-1H-pyrazole-3-carboxylate;-   Ethyl    1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate;-   Ethyl    1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(trimethylsilyl)-1H-pyrazole-5-carboxylate;-   Ethyl    1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(trimethylsilyl)-1H-pyrazole-5-carboxylate;-   Ethyl    3-(pyridin-2-yl)-1-(pyridin-3-ylmethyl)-1H-pyrazole-5-carboxylate;-   Ethyl    1-(3,4,5-trimethoxybenzyl)-3-(trimethylsilyl)-1H-pyrazole-5-carboxylate;-   Ethyl    1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate;-   Ethyl    1-(2-oxo-2-(pyridin-3-yl)ethyl)-5-(trimethylsilyl)-1H-pyrazole-3-carboxylate;-   Ethyl    1-(2-oxo-2-(pyridin-3-yl)ethyl)-3-(trimethylsilyl)-1H-pyrazole-5-carboxylate;-   Ethyl    1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate;    or-   Ethyl    1-((1H-indol-3-yl)methyl)-3-isopropyl-1H-pyrazole-5-carboxylate.

In a further embodiment of the invention, provided is a pharmaceuticalcomposition, comprising a therapeutically effective amount of a compoundaccording to formula (I) or a pharmaceutically acceptable salt thereofand a pharmaceutically acceptable carrier.

In a further embodiment of the invention, provided is a method fortreating a degenerative disease or disorder, comprising the step ofadministering a therapeutically effective amount of a compound accordingto formula (I) or a pharmaceutically acceptable salt thereof and apharmaceutically acceptable carrier to a patient in need thereof. Thedegenerative diseases and disorders include, for example, retinitispigmentosa.

In another embodiment of the invention, provided is a method of treatinga retinal degenerative disease in a subject in need thereof, comprisingadministering to said subject a therapeutically effective amount of acompound or a pharmaceutically acceptable salt thereof according toformula (I).

In a yet further embodiment of the invention, provided is a method forpreventing calcium-induced or oxidant-induced mitochondrial damagepreventing or loss of mitochondrial respiratory capacity in a cellsusceptible thereof wherein the calcium-induced or oxidant-inducedmitochondrial damage or loss of mitochondrial respiratory capacitycomprises excess of cGMP that increases the number of cGMP-gated cationchannels in an open configuration, allowing an influx of Ca2+ into thecell, said method comprising contacting the cell with an effectiveamount of a compound or a pharmaceutically acceptable salt thereofaccording to formula (I).

In the practice of the method of the present invention, an effectiveamount of any one of the compounds of this invention or a combination ofany of the compounds of this invention or a pharmaceutically acceptablesalt thereof, is administered via any of the usual and acceptablemethods known in the art, either singly or in combination. The compoundsor compositions can thus be administered, for example, ocularly, orally(e.g., buccal cavity), sublingually, parenterally (e.g.,intramuscularly, intravenously, or subcutaneously), rectally (e.g., bysuppositories or washings), transdermally (e.g., skin electroporation)or by inhalation (e.g., by aerosol), and in the form or solid, liquid orgaseous dosages, including tablets and suspensions. The administrationcan be conducted in a single unit dosage form with continuous therapy orin a single dose therapy ad libitum. The therapeutic composition canalso be in the form of an oil emulsion or dispersion in conjunction witha lipophilic salt such as pamoic acid, or in the form of a biodegradablesustained-release composition for subcutaneous or intramuscularadministration.

Useful pharmaceutical carriers for the preparation of the compositionshereof, can be solids, liquids or gases. Thus, the compositions can takethe form of tablets, pills, capsules, suppositories, powders,enterically coated or other protected formulations (e.g. binding onion-exchange resins or packaging in lipid-protein vesicles), sustainedrelease formulations, solutions, suspensions, elixirs, aerosols, and thelike. The carrier can be selected from the various oils including thoseof petroleum, animal, vegetable or synthetic origin, e.g., peanut oil,soybean oil, mineral oil, sesame oil, and the like. Water, saline,aqueous dextrose, and glycols are preferred liquid carriers,particularly (when isotonic with the blood) for injectable solutions.For example, formulations for intravenous administration comprisesterile aqueous solutions of the active ingredient(s) which are preparedby dissolving solid active ingredient(s) in water to produce an aqueoussolution, and rendering the solution sterile. Suitable pharmaceuticalexcipients include starch, cellulose, talc, glucose, lactose, talc,gelatin, malt, rice, flour, chalk, silica, magnesium stearate, sodiumstearate, glycerol monostearate, sodium chloride, dried skim milk,glycerol, propylene glycol, water, ethanol, and the like. Thecompositions may be subjected to conventional pharmaceutical additivessuch as preservatives, stabilizing agents, wetting or emulsifyingagents, salts for adjusting osmotic pressure, buffers and the like.Suitable pharmaceutical carriers and their formulation are described inRemington's Pharmaceutical Sciences by E. W. Martin. Such compositionswill, in any event, contain an effective amount of the active compoundtogether with a suitable carrier so as to prepare the proper dosage formfor proper administration to the recipient.

The dose of a compound of the present invention depends on a number offactors, such as, for example, the manner of administration, the age andthe body weight of the subject, and the condition of the subject to betreated, and ultimately will be decided by the attending physician orveterinarian. Such an amount of the active compound as determined by theattending physician or veterinarian is referred to herein, and in theclaims, as a “therapeutically effective amount”. For example, the doseof a compound of the present invention is typically in the range ofabout 1 to about 1000 mg per day. Preferably, the therapeuticallyeffective amount is in an amount of from about 1 mg to about 500 mg perday.

It will be appreciated, that the compounds of general formula I in thisinvention may be derivatized at functional groups to provide derivativeswhich are capable of conversion back to the parent compound in vivo.Physiologically acceptable and metabolically labile derivatives, whichare capable of producing the parent compounds of general formula I invivo are also within the scope of this invention.

Compounds of the present invention can be prepared beginning withcommercially available starting materials and utilizing generalsynthetic techniques and procedures known to those skilled in the art.Chemicals may be purchased from companies such as for example Aldrich,Argonaut Technologies, VWR and Lancaster. Chromatography supplies andequipment may be purchased from such companies as for example AnaLogix,Inc, Burlington, Wis.; Biotage AB, Charlottesville, Va.; AnalyticalSales and Services, Inc., Pompton Plains, N.J.; Teledyne Isco, Lincoln,Nebr.; VWR International, Bridgeport, N.J.; Varian Inc., Palo Alto,Calif., and Multigram II Mettler Toledo Instrument Newark, Del. Biotage,ISCO and Analogix columns are pre-packed silica gel columns used instandard chromatography.

The compounds of formula I can be prepared according to the followingscheme:

As seen in Scheme 1, compounds of formula I and II (collectively“formula (I)”) may be made using intermediate i. Intermediate i may bemade from reacting an acetylene where R₁ can be, for example, aryl,phenyl, 2-pyridyl, or 3-pyridyl, methyl, tert-butyl, trimethyl silyl,trialkyl silyl, dialkylphenylsilyl, diphenylalkylsilyl, ortriphenylsilyl with the appropriately commercially available diazoethylacetate (purchased from Aldrich) at the appropriate temperature (such as95° C.) for the appropriate time (such as 24 hours) (Cheung, K. M. J.;Reynisson, J.; McDonald, E. Tetrahedron Lett. 2010, 51 5915-5918).Formation of compounds of formula I may then be made by reactingintermediates of formula i with a base such as LiHMDS, KHMDS, NaHMDS,LDA, BuLi, t-BuMgC1, any alkyl lithium, any aryl lithium, any alkylGrignard, or any aryl Grignard, that may or may not be in the presenceof 18-crown-6, or compounds analogous thereto, in a solvent such as DMF,THF, or 1,4 dioxane at the appropriate temperature with any commerciallyavailable R₂—X to afford compounds of formulation I or II as either amixture, or exclusive. R₂ and R_(2′), independently of each other, maybe, for example, benzyl, aryl, aryl keto,2,4-dimethoxyphenyl)-2-oxoethyl, (2,5-dimethoxyphenyl)-2-oxoethyl,pyridin-3-ylmethyl, 3,4,5-trimethoxybenzyl, 2-oxo-2-(pyridin-3-yl)ethyl, -(1H-indol-3-yl)methyl. X may be any halogen such as chlorine,bromine, or iodine.

The invention will now be further described in the Examples below, whichare intended as an illustration only and do not limit the scope of theinvention.

EXAMPLES I. Preparation of Certain Intermediates of the Invention Ethyl3-(trimethylsilyl)-1H-pyrazole-5-carboxylate

To a flame dried sealed tube equipped with a stir bar that was cooledunder argon was added trimethylsilylacetylene (1.0 mL, 9.56 mmol) andethyldiazoacetate (1.5 mL, 9.6 mmol). The tube was then sealed andheated to 95° C. over night. The next day the reaction was cooled toroom temperature and the resulting mixture diluted with hexanes. It wasthen filtered. The precipitate was then washed with hexanes twice. Itwas then used without any further purification.

Ethyl 3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate

Was prepared in a similar way as ethyl3-(trimethylsilyl)-1H-pyrazole-5-carboxylate using diazoethyl acetate(Purchased from Aldrich) and 2-ethynyl-pyridine (Purchased fromAldrich).

Ethyl 3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate

Was prepared in a similar way to ethyl3-(trimethylsilyl)-1H-pyrazole-5-carboxylate using diazoethyl acetate(Purchased from Aldrich) and 2-ethynyl-pyridine (Purchased from Aldrich)

1-Benzoyl-1H-indol-3-yl)methyl benzoate

To an oven dried flask that cooled under argon was added the(1H-indol-3-yl)methanol (1.0 grams, 6.8 mmol, 0.1M in drydichloromethane, purchased from Fisher Scientific, stored over 4angstrom molecular sieves) and DMAP (0.083 grams, 0.68 mmol). Whilestirring at 0° C., triethyl amine (2.0 mL, 14.3 mmol, purchased fromFisher Scientific) was added followed by benzoyl chloride (0.96 mL, 8.2mmol, purchased from Fisher Scientific). Once the reaction was completeit was diluted with water, and the organic layer removed. The aqueouslayer was then washed with dichloromethane twice and the organicmaterial combined. The organic material was dried with sodium sulfate,filtered, and concentrated. Purification using a Teledyne ISCO on silicasupport (hexanes/ethyl acetate gradient) afforded the desired1-benzoyl-1H-indol-3-yl)methyl benzoate. 42% yield. 1H-NMR δ8.42 (d,1H), 8.02 (dd, 2H), 7.76 (m, 3H), 7.62 (dd, 1H), 7.54 (m, 3H), 7.47-7.37(m, 5H), 5.50 (s, 2H).

Ethyl 3-isopropyl-1H-pyrazole-5-carboxylate

To a clean round bottom flask equipped with a stir bar, Dean-Stark trap,and reflux condenser was added 3-isopropyl-1H-pyrazole-5-carboxylic acid(1 gram, 6.49 mmol, purchased from Fisher Scientific). 30 mL of ethanol(95%, purchased from Fisher Scientific) was added followed by 30 mL ofbenzene. 43 μL of acetyl chloride was then added and the solutionrefluxed over the three days. The solvent was then removed using a Buchirotoevaporator. The residue was then taken up in ethyl acetate andwashed with NaHCO3 (saturated). It was then dried with sodium sulfate,filtered and concentrated. Purification using a Teledyne ISCO silicachromatography (hexanes/ethyl acetate gradient) afforded the desiredester. Yield, 85% 1H-NMR δ 6.7 (s, 1H), 5.1 (bs, NH), 4.41 (q, 2H), 3.13(septet, 1H), 1.38 (t, 3H), 1.33 (d, 6H).

II. Preparation of Certain Embodiments of the Invention Examples 1 and 2Ethyl1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-2-yl)-1H-pyrazole-5-carboxylateand Ethyl1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-5-(pyridin-2-yl)-1H-pyrazole-3-carboxylate

To an oven dried flask equipped with a stir bar cooled under argon wasadded ethyl 3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate (0.03 grams, 0.14mmol, 0.1M in 1,4 dioxane (anhydrous Sure Seal, purchased fromAldrich)). While stirring at room temperature, a solution of KHMDS (0.17mL, 0.15 mmol, 0.87 M in toluene, purchased from Fisher Scientific) wasadded slowly. In a separate oven dried flask cooled under argon wasadded 2-bromo-1-(2,4-dimethoxyphenyl)ethanone (0.04 grams, 0.154 mmol,0.1M in 1,4 dioxane (anhydrous Sure Seal, purchased from Aldrich)).

After stirring for one hour at room temperature the solution ofdimethoxyphenylethanone was added to the ethyl3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate and the reaction continued tostir at room temperature over night. The next day, the reaction wasdiluted with 0.1 M HCl and ethyl acetate. The organic material wasextracted. The aqueous layer was salted out with sodium chloride andwashed twice with ethyl acetate. The combined organic material was thendried with sodium sulfate, filtered and concentrated. Purification usinga Teledyne ISCO on a silica support (hexanes, ethyl acetate gradient)affords the two regioisomers in a 1:1 ratio. Further purification can bedone using a Teledyne ISCO C18 reverse phase column using water with0.1% formic acid, acetonitrile gradient. Combined yield, 60%.1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate1H-NMR □ 8.34 (ddd, 1H), 7.81 (d, 1H), 7.69 (m, 2H), 7.29 (s, 1H), 7.14(m, 1H), 6.54 (dd, 1H), 6.51 (d, 1H), 6.23 (s, 2H), 4.4 (q, 2H), 3.97(s, 3H), 3.88 (s, 3H), 1.43 (t, 3H). calculated mass for C21H21N3O5,395.15, observed, 396.2 (M+1).

1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-5-(pyridine-2-yl)-1H-pyrazole-3-carboxylate.1H-NMR δ8.33 (dd, 1H), 7.81 (d, 1H), 7.69 (m, 2H), 7.28 (s, 1H), 7.14(t, 1H), 6.54 (dd, 1H), 6.50 (d, 1H), 6.22 (s, 2H), 4.44 (d, 2H), 3.96(s, 3H), 3.88 (s, 3H), 1.42 (t, 3H). Calculated mass for C21H21N3O5,395.15, observed, 418.1 (M+Na).

Example 3 Ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate

To a flame dried flask equipped with a stirbar cooled under argon wasadded ethyl 3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate (0.02 grams,0.092 mmol, 0.1 M in THF). While stirring at room temperature NaH(0.0055 grams, 1.8 mmol, 60% in mineral oil, purchase from Aldrich) wasadded. After fifteen minutes 2-bromo-1-(2,5-dimethoxyphenyl)ethanone(0.047 grams, 0.14 mmol, purchased from Aldrich) was added as a solid.The reaction was stirred overnight. The next day, the reaction wasquenched with 0.1M HCl and the organic material extracted using ethylacetate. The aqueous layer was salted out using sodium chloride andwashed twice with ethyl acetate. The combined organic material was thendried with sodium sulfate, filtered, and concentrated. Purification wasdone on preparative thin layer chromatography using hexanes/ethylacetate (1:2) to afford the desired compound. Yield, 10%. 1H-NMR δ9.07(bs, 1H), 8.58 (bs, 1H), 8.13 (d, 1H), 7.47 (d, 1H), 7.31 (bs, 1H), 7.29(s, 1H), 7.14 (dd, 1H), 7.00 (d, 1H), 6.03 (s, 2H), 4.32 (q, 2H), 3.99(s, 3H), 3.79 (s, 3H), 1.35 (t, 3H). Calculated mass for C21H21N3O5,395.15, observed, 396.3 (M+1).

Example 4 Ethyl1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(trimethylsilyl)-1H-pyrazole-5-carboxylate

This compound was prepared in a similar method to ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylateusing ethyl 3-(trimethyl silyl)-1H-pyrazole-5-carboxylate and2-bromo-1-(2,4-dimethoxyphenyl)ethanone (purchased from Aldrich). 1H-NMRδ 9.92 (d, 1H), 7.02 (s, 1H), 6.57 (dd, 1H), 6.51 (d, 1H), 5.67 (s, 2H),4.39 (quartet, 2H), 3.98 (s, 3H), 3.89 (s, 3H), 1.38 (t, 3H), 0.27 (s,9H).

Example 5 Ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(trimethylsilyl)-1H-pyrazole-5-carboxylate

This compound was prepared in a similar method to ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylateusing ethyl 3-(trimethylsilyl)-1H-pyrazole-5-carboxylate and2-bromo-1-(2,5-dimethoxyphenyl)ethanone (Purchased from Aldrich). 1H-NMRδ 7.40 (d, 1H), 7.12 (dd, 1H), 7.02 (dd, 1H), 6.96 (d, 1H), 5.72 (s,2H), 4.40 (q, 2H), 3.96 (s, 3H), 3.78 (s, 3H), 1.39 (t, 3H), 0.28 (s,9H).

Example 6 Ethyl3-(pyridin-2-yl)-1-(pyridin-3-ylmethyl)-1H-pyrazole-5-carboxylate

To a flame dried flask equipped with a stir bar cooled under argon wasadded ethyl 3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate (0.02 grams,0.092 mmol), 3-bromo-methylene-pyridine.HBr (0.0466 grams, 0.18 mmol,purchased from Aldrich) and 1 mL of THF (anhydrous, Sure Seal purchasedfrom Aldrich). While stirring at room temperature sodium hydride (0.011grams, 0.28 mmol, 60% in mineral oil, purchased from Aldrich), was addedand the reaction continued to stir over night at room temperature. Thenext day, the reaction was quenched with 0.1M HCl, and the organicmaterial extracted using ethyl acetate. The aqueous layer was thensalted out using sodium chloride, and washed twice with ethyl acetate.The combined organic material was then dried with sodium sulfate,filtered, and concentrated. Purification was done on preparative thinlayer chromatography using hexanes/ethyl acetate (1:2) to afford thedesired compound. Yield, 9.4%. ¹H-NMR δ 8.67 (m, 2H), 8.56 (m, 1H), 8.00(d, 1H), 7.80-7.72 (m, 2H), 7.56 (m, 1H), 7.35-7.24 (m, 2H), 5.88 (s,2H), 4.34 (q, 2H), 1.36 (t, 3H).

Example 7 Ethyl1-(3,4,5-trimethoxybenzyl)-3-(trimethylsilyl)-1H-pyrazole-5-carboxylate

This compound was made in an analogous fashion to ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylateusing ethyl 3-(trimethylsilyl)-1H-pyrazole-5-carboxylate and3,4,5-trimethoxy benzyl chloride (purchased from Aldrich). 1H-NMR δ 6.98(s, 1H), 6.21 (s, 2H), 5.44 (s, 2H), 4.41 (q, 2H), 3.81 (s, 3H), 3.76(s, 6H), 1.40 (t, 3H), 0.22 (s, 9). Calculated mass for C19H28N2O5Si,392.18, observed, 393.0 (M+1), 415.1 (M+Na).

Example 8 Ethyl1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate

This compound was made in an analogous fashion to ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylateusing ethyl 3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate and2-bromo-1-(2,4-dimethoxyphenyl)ethanone (purchased from Aldrich). 1H-NMRδ 9.08 (s, 1H), 8.58 (d, 1H), 8.14 (d, 1H), 8.00 (d, 1H), 7.33 (m,11-1), 7.27 (s, 1H), 6.58 (dd, 1H), 6.49 (d, 1H), 5.98 (s, 2H), 4.31 (q,2H), 4.00, (s, 3H), 3.88 (s, 3H), 1.34 (t, 3H). Calculated mass forC21H21N3O5, 395.15, observed, 396.2 (M+1).

Example 9 Ethyl1-(2-oxo-2-(pyridin-3-yl)ethyl)-5-(trimethylsilyl)-1H-pyrazole-3-carboxylate

This compound was made in an analogous fashion to ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylateusing ethyl 3-(trimethylsilyl)-1H-pyrazole-5-carboxylate and2-bromo-1-(pyridin-3-yl)ethanone.HBr (purchased from Aldrich). 1H-NMR δ9.17 (d, 1H), 8.88 (dd, 1H), 8.24 (ddd, 1H), 7.50 (td, 1H), 7.03 (s,1H), 5.73 (s, 2H), 4.40 (q, 2H), 1.39 (t, 3H), 0.28 (s, 9H). Calculatedmass for C16H21N3O3Si, 331.14, observed, 332.0 (M+1).

Example 10 Ethyl1-(2-oxo-2-(pyridin-3-yl)ethyl)-3-(trimethylsilyl)-1H-pyrazole-5-carboxylate

This compound was made in an analogous fashion to ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylateusing ethyl 3-(trimethylsilyl)-1H-pyrazole-5-carboxylate and2-bromo-1-(pyridin-3-yl)ethanone.HBr (purchased from Aldrich). 1H-NMR δ9.20 (d, 1H), 8.84 (dd, 1H), 8.25 (dt, 1H), 7.46 (dd, 1H), 7.07 (s, 1H),6.05 (s, 2H), 4.26 (q, 2H), 1.31 (t, 3H), 0.32 (s, 9H). Calculated massfor C16H21N3O3Si, 331.14, observed, 332.0 (M+1).

Example 11 Ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate

This compound was made in an analogous fashion to ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylateusing ethyl 3-(trimethylsilyl)-1H-pyrazole-5-carboxylate and2-bromo-1-(2,5-dimethoxyphenyl)ethanone (purchased from Aldrich). 1H-NMRδ 8.65 (dt, 1H), 7.94 (d, 1H), 7.72 (dd, 1H), 7.57 (s, 1H), 7.47 (d,1H), 7.22 (dd, 1H), 7.13 (dd, 1H), 6.98 (s, 1H), 6.05 (s, 2H), 4.28 (q,2H), 3.98 (s, 3H), 3.78 (s, 3H), 1.33 (t, 3H). Calculated mass forC21H21N3O5, 395.15, observed, 396.2 (M+1).

Example 12 Ethyl1-((1H-indol-3-yl)methyl)-3-isopropyl-1H-pyrazole-5-carboxylate

To an oven dried flask cooled under argon equipped with a stir bar wasadded ethyl 3-isopropyl-1H-pyrazole-5-carboxylate (0.044 grams, 0.24mmol, 0.1 in anhydrous 1,4 dioxane). While stirring at room temperatureKHMDS (0.3 mL, 0.261 mmol, 0.87 M in toluene) was added. After stirringfor 45 minutes, a solution of (1-benzoyl-1H-indol-3-yl)methyl benzoate(0.02 grams, 0.056 mmol, in 1.0 mL of anhydrous 1,4 dioxane) was addedand the reaction mixture stirred over night. The next day, the reactionwas quenched with 0.1M HCl and the organic material extracted with ethylacetate. The aqueous solution was then salted out using sodium chlorideand washed twice with ethyl acetate. The combined organic material wasthen dried with sodium sulfate, filtered, and concentrated. Purificationusing a Teledyne ISCO chromatography on silica gel (hexanes/ethylacetate gradient) followed by a Teledyne ISCO chromatography using C18reverse phase (water with 0.1% formic acid, acetonitrile gradient)afforded the desired compound. Yield, 63%. ¹H NMR δ 8.29 (bs, 1H), 7.58(d, 1H), 7.36 (d, 1H), 7.20 (t, 1H), 7.11 (t, 1H), 6.96 (s, 1H), 6.62(s, 1H), 5.58 (s, 2H), 4.41 (q, 2H), 3.03 (m, 1H), 1.39 (t, 3H), 1.16(d, 6H). Calculated mass for C18H21N3O2, 311.16. Observed 334.1 (M+1).

Example 13 Biological Assays of Certain Compounds of the Invention

The compounds of the invention were tested in various biological assays.The results of these assays indicated that the compounds of theinvention ameliorated dysregulated bioenergetics and are, thus, usefulfor treatment of degenerative diseases and disorders, such as retinaldamage.

MTT Assay

The compound 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) is a tetrazolium ion that is reduced to a blue formazandye via several families of NAD(P)H-dependent oxidoreductases. Formationof the formazan dye from MTT or other related tetrazolium dyes arecommonly used as a viability assay even though, in fact, the assay is ametabolic capacity assay. While it is true that dead cells cannotproduce NAD(P)H, very sick cells in the throes of death can exhibitextremely high levels of metabolic capacity as they attempt to overcomestress and it is well known that the MTT and related assays report onthe ability of cells to produce reducing equivalents, and not live-deadratios (Sumantran 2011). As shown below, it was found that the MTT assaywas a useful metabolic assay when linked to more specific bioenergeticassays.

In this assay, 661W or C6 cells were maintained in DMEM supplementedwith 10% FBS. 100 μL of 70,000 cells/mL cells were seeded into each wellof 96 well plates using DMEM supplemented with 5% FBS. Cells were thenallowed to grow to confluency for 48 hours. The compounds of theinvention were added in 2 μL media and calcium-ionophore A23187 was thenadded in 1 μL for a final concentration of 1 μM and after 24 h, 20 uL of2 μg/mL MTT were added to each well and the cells were incubated foranother 4 h after which 100 μL of 1% SDS in 0.01 M aqueous HCl wereadded to each well and the plates were incubated overnight. Absorbancewas measured at 640 and 570 nM (background correction). The 1 μMionophore A23187 caused about 50% loss in MTT signal at 24 h. Protectionwas calculated as the increase in absorbance of treatment groupsnormalized to the vehicle control. As shown in Table 1, the compounds ofthe invention gave significant protection at low concentrations:

TABLE 1 MTT Protection Example (%, concentration) 1 10.4%, 800 pM 273.9%, 800 pM 3 43.5%, 1 nM 4   87%, 10 nM 5 76.6%, 10 nM 6 73.5%, 10 nM7   40%, 10 nM 8 75.6%, 1 nM 9   59%, 1 nM 10 47.8%, 1 nM 11 28.3%, 1 μM12 59.4%, 1 nMXF FCCP-Uncoupled Oxygen Consumption Rate Assay

The XF FCCP-uncoupled oxygen consumption rate assay assessesmitochondrial capacity by measuring cellular respiration. It was shownthat the maximum FCCP-uncoupled oxygen consumption rate (OCR) was a goodestimate of maximal mitochondrial capacity (Beeson 2010) and that IBMXtreatment of 661W or C6 cells for 24 h caused a loss in maximaluncoupled OCR (Perron 2013). Thus, cells were pretreated with theexemplified compounds for 1 h, added 600 μM IBMX and then measured theuncoupled rate after 24 h. The OCR measurements were performed using aSeahorse Bioscience XF instrument (Seahorse Bioscience, Billerica, Md.),as previously published (Perron 2013). O₂ leakage through the plasticsides and bottom of the plate was accounted for using the AKOS algorithmin the XF software package. Cells were plated on 96- or 24-well customplates designed for use in the XF and grown to confluency in DMEM+5% FBS(48 h). The medium was then replaced with DMEM+1% FBS for 24 h, alongwith any treatments. The IBMX treatment alone typically caused about a50% decrease in the uncoupled rate and protection was calculated as theincrease in absorbance of treatment groups normalized to the vehiclecontrol. As shown in Table 2, below, the compounds of the invention gavesignificant protection in the concentration ranges that gave maximalprotection in the MTT assay:

TABLE 2 XF FCCP OCR Example No. (% Ctrl, concentration) 1 58% @ 1 uM 262% @ 100 nM 3 Not Tested 4 67% @ 10 nM 5 72% @ 10 nM 6 75% @ 10 nM 864% @ 100 nM 9 67% @ 100 nMRetinal Degeneration Assay

The in vitro data demonstrated that the compounds of the inventionmitigated oxidative- and calcium-induced loss of mitochondrial metaboliccapacity. It was reasoned that the compounds' activities would enablethem to protect against loss of photoreceptors in retinal degenerativeanimal models. As a translational bridge between the cell line-basedassays and in vivo animal studies, mouse retina organ cultures wereutilized. These retinal explants were a powerful ex vivo screening tool,which allowed the testing of photoreceptor cell survival within theretinal network, and the effects of a specific compound were tested likein an in vitro system, without systemic interference. In this assay, therd1 mouse was utilized. The genotype of the rd1 mouse has a mutation inthe β-subunit of the phosphodiesterase gene. This mutation resulted inhigh levels of cGMP, leaving an increased number of the cGMP-gatedchannels in the open state, allowing intracellular calcium to rise totoxic levels and rapid rod degeneration. The genetic deficit and theretinal pathology were very similar to that observed in the patientswith βPDE-dependent RP. In these mice, rod photoreceptor degenerationstarted after postnatal day 10 (P10), progressing rapidly, such that atP21, only 1-2 rows of photoreceptor remained, mainly representing cones.Finally, the rd1 mouse retina was amenable to culturing, replicatingboth retinal development and degeneration, following the same timecourse as in vivo. The retinal explants were cultured for 11 days exvivo. Explants were treated with compounds of the invention. Additiveswere replaced with fresh medium in every alternate day. At the end ofthe experiments, tissues were fixed, sectioned and stained with 0.1%toluidine and the numbers of rows of photoreceptors remaining in theouter nuclear layer (ONL) were counted. Rd1 explants treated withvehicle only were found to contain 1.2±0.19 cells in the ONL. This wasin contrast to cultures treated with the compounds of the invention thatshowed significant protection (Table 3 below):

TABLE 3 Rd1 vehicle rd1 protection Example (# of (concentration, No.rows) # of rows) 2 1.3 20 nM 4.46 4 1.16 10 nM 3.2 Light Model Assay

The light model assay is generally accepted as a model of age relatedmacular degeneration (AMD). Light as an environmental factor has beenshown to be toxic to rod photoreceptors if the retina was exposed tohigh light levels over a long period of time; and oxidative stress hasbeen implicated as the main trigger for cell death. In particular,oxidative damage has been detected by immunohistochemistry, detectingthe presence of oxidized and tyrosine-phosphorylated proteins as well asthe upregulation of endogenous antioxidants such as thioredoxin andglutathione peroxidase. Likewise, exogenous antioxidants have been foundto protect the rodent retina from light damage. Additional indirectevidence for the involvement of oxidative stress in photoreceptordegeneration has been provided by treatment of photodamaged retinas withantioxidants such as dimethylthiourea, or the treatment ofN-methyl-N-nitrosourea (MNU)-challenged rats with the antioxidant DHA.

The light model assay was used to further test the therapeutic potentialof the compounds of the invention. Photoreceptors from albino animalsare very sensitive to constant light, lacking the RPE pigment to protectthem. Thus, Balb/c mice were exposed to continuous light for 7 days,which caused loss of about 50% of the photoreceptor cells as measuredvia histology. To test the potential therapeutic efficacy, eyedrops wereformulated in 0.1% Bij 35 in 9% saline, applied once or twice dailythroughout the period of light exposure, and assessed their effect onthe light-induced degeneration of photoreceptor cells morphologicallyand electrophysiologically, 10 days after the onset of the CL exposure.In control BALB/c mice, constant light resulted in the elimination of˜50% of the photoreceptors (average retina score: 4.3±0.25 rows ofphotoreceptors), whereas the mice treated with compound eyedrops onceper day retained significantly more photoreceptors cells (Table 4).

TABLE 4 LD Protection (concentration, Example LD vehicle (# of rows) #of rows) 4 4.95 125 μM 6.42

As seen above, the compounds of the invention mitigate oxidative- andcalcium-mediated loss of mitochondrial capacity in cell lines andprotect photoreceptors from cell death in several models of retinaldegeneration.

REFERENCES

-   Acosta M L, Fletcher E L, Azizoglu S, Foster L E, Farber D B,    Kalloniatis M: Early markers of retinal degeneration in rd/rd mice.    Mol Vis 2005, 11:717-728.-   Acosta M L, Shin Y S, Ready S, Fletcher E L, Christie D L,    Kalloniatis M. Retinal metabolic state of the proline-23-histidine    rat model of retinitis pigmentosa. Am J Physiol Cell Physiol. 2010    March; 298(3):C764-74. doi: 10.1152/ajpcell.00253.2009. Epub 2009    Dec. 23. PubMed PMID: 20032515.-   Bandyopadhyay M, Rohrer B. Photoreceptor structure and function is    maintained in organotypic cultures of mouse retinas. Mol Vis. 2010    Jun. 26; 16:1178-85. PubMed PMID: 20664685; PubMed Central PMCID:    PMC2901185.-   Barot M, Gokulgandhi M R, Mitra A K. Mitochondrial dysfunction in    retinal diseases. Curr Eye Res. 2011 December; 36(12):1069-77. doi:    10.3109/02713683.2011.607536. Epub 2011 Oct. 6. Review. PubMed PMID:    21978133.-   Beal D M, Jones L H. Molecular scaffolds using multiple orthogonal    conjugations: applications in chemical biology and drug discovery.    Angew Chem Int Ed Engl. 2012 Jun. 25; 51(26):6320-6. doi:    10.1002/anie.201200002. Epub 2012 Apr. 19. Review. PubMed PMID:    22517597.-   Beeson C C, Beeson G C, Schnellmann R G. A high-throughput    respirometric assay for mitochondrial biogenesis and toxicity. Anal    Biochem. 2010 Sep. 1; 404(1):75-81. doi: 10.1016/j.ab.2010.04.040.    Epub 2010 May 11. PubMed PMID: 20465991; PubMed Central PMCID:    PMC2900494.-   Booij J C, van Soest S, Swagemakers S M, Essing A H, Verkerk A J,    van der Spek P J, Gorgels T G, Bergen A A. Functional annotation of    the human retinal pigment epithelium transcriptome. BMC Genomics.    2009 Apr. 20; 10:164. doi: 10.1186/1471-2164-10-164. PubMed PMID:    19379482; PubMed Central PMCID: PMC2679759.-   Bruce J E. In vivo protein complex topologies: sights through a    cross-linking lens. Proteomics. 2012 May; 12(10):1565-75. doi:    10.1002/pmic.201100516. Review. PubMed PMID: 22610688.-   Catoire M, Mensink M, Boekschoten M V, Hangelbroek R, Müller M,    Schrauwen P, Kersten S. Pronounced effects of acute endurance    exercise on gene expression in resting and exercising human skeletal    muscle. PLoS One. 2012; 7(11):e51066. doi:    10.1371/journal.pone.0051066. Epub 2012 Nov. 30. PubMed PMID:    23226462; PubMed Central PMCID: PMC3511348.-   Cavalier-Smith T, Chao E E. Phylogeny of choanozoa, apusozoa, and    other protozoa and early eukaryote megaevolution. J Mol Evol. 2003    May; 56(5):540-63. PubMed PMID: 12698292.-   Cazares L H, Troyer D A, Wang B, Drake R R, Semmes O J. MALDI tissue    imaging: from biomarker discovery to clinical applications. Anal    Bioanal Chem. 2011 July; 401(1):17-27. doi:    10.1007/s00216-011-5003-6. Epub 2011 May 4. Review. PubMed PMID:    21541816.-   Chaurand P, Cornett D S, Caprioli R M. Molecular imaging of thin    mammalian tissue sections by mass spectrometry. Curr Opin    Biotechnol. 2006 August; 17(4):431-6. Epub 2006 Jun. 16. Review.    PubMed PMID: 16781865.-   Chen Y A, Almeida J S, Richards A J, Müller P, Carroll R J,    Rohrer B. A nonparametric approach to detect nonlinear correlation    in gene expression. J Comput Graph Stat. 2010 Sep. 1; 19(3):552-568.    PubMed PMID: 20877445; PubMed Central PMCID: PMC2945392.-   Copple I M. The Keap1-Nrf2 cell defense pathway—a promising    therapeutic target Adv Pharmacol. 2012; 63:43-79. doi:    10.1016/B978-O-12-398339-8.00002-1. Review. PubMed PMID: 22776639.-   Court F A, Coleman M P. Mitochondria as a central sensor for axonal    degenerative stimuli. Trends Neurosci. 2012 June; 35(6):364-72. doi:    10.1016/j.tins.2012.04.001. Epub 2012 May 11. Review. PubMed PMID:    22578891.-   Dai C, Cazares L H, Wang L, Chu Y, Wang S L, Troyer D A, Semmes O J,    Drake R R, Wang B. Using boronolectin in MALDI-MS imaging for the    histological analysis of cancer tissue expressing the sialyl Lewis X    antigen. Chem Commun (Camb). 2011 Oct. 7; 47(37):10338-40. doi:    10.1039/cicc11814e. Epub 2011 Aug. 19. PubMed PMID: 21853197.-   Daiger S P, Sullivan L S, Bowne S J, Birch D G, Heckenlively J R,    Pierce E A, Weinstock G M. Targeted high-throughput DNA sequencing    for gene discovery in retinitis pigmentosa. Adv Exp Med. Biol. 2010;    664:325-31. doi: 10.1007/978-1-4419-1399-9_(—)37. PubMed PMID:    20238032; PubMed Central PMCID: PMC2909649.-   De Jesús-Cortés H, Xu P, Drawbridge J, Estill S J, Huntington P,    Tran S, Britt J, Tesla R, Morlock L, Naidoo J, Melito L M, Wang G,    Williams N S, Ready J M, McKnight S L, Pieper A A. Neuroprotective    efficacy of aminopropyl carbazoles in a mouse model of Parkinson    disease. Proc Natl Acad Sci USA. 2012 Oct. 16; 109(42):17010-5. doi:    10.1073/pnas.1213956109. Epub 2012 Oct. 1. PubMed PMID: 23027934;    PubMed Central PMCID: PMC3479520.-   Demos C, Bandyopadhyay M, Rohrer B. Identification of candidate    genes for human retinal degeneration loci using differentially    expressed genes from mouse photoreceptor dystrophy models. Mol Vis.    2008 Sep. 5; 14:1639-49. PubMed PMID: 18776951; PubMed Central    PMCID: PMC2529471.-   Dong S Q, Xu H Z, Xia X B, Wang S, Zhang L X, Liu S Z. Activation of    the ERK 1/2 and STAT3 signaling pathways is required for 661W cell    survival following oxidant injury. Int J. Ophthalmol. 2012;    5(2):138-42. doi: 10.3980/j.issn.2222-3959.2012.02.04. Epub 2012    Apr. 18. PubMed PMID: 22762037; PubMed Central PMCID: PMC3359025.-   Egger A, Samardzija M, Sothilingam V, Tanimoto N, Lange C, Salatino    S, Fang L, Garcia-Gamido M, Beck S, Okoniewski M J, Neutzner A,    Seeliger M W, Grimm C, Handschin C. PGC-1α determines light damage    susceptibility of the murine retina. PLoS One. 2012; 7(2):e31272.    doi: 10.1371/journal.pone.0031272. Epub 2012 Feb. 13. PubMed PMID:    22348062; PubMed Central PMCID: PMC3278422.-   Estrada-Cuzcano A, Roepman R, Cremers F P, den Hollander A I, Mans    D A. Non-syndromic retinal ciliopathies: translating gene discovery    into therapy. Hum Mol Genet. 2012 Oct. 15; 21(R1):R111-24. Epub 2012    Jul. 26. PubMed PMID: 22843501.-   Falk M J, Zhang Q, Nakamaru-Ogiso E, Kannabiran C, Fonseca-Kelly Z,    Chakarova C, Audo I, Mackay D S, Zeitz C, Borman A D, Staniszewska    M, Shukla R, Palavalli L, Mohand-Said S, Waseem N H, Jalali S, Perin    J C, Place E, Ostrovsky J, Xiao R, Bhattacharya S S, Consugar M,    Webster A R, Sahel J A, Moore A T, Berson E L, Liu Q, Gai X, Pierce    E A. NMNAT1 mutations cause Leber congenital amaurosis. Nat. Genet.    2012 September; 44(9):1040-5. doi: 10.1038/ng.2361. Epub 2012    Jul. 29. PubMed PMID: 22842227; PubMed Central PMCID: PMC3454532.-   Farber D B, Lolley R N: Cyclic guanosine monophosphate: elevation in    degenerating photoreceptor cells of the C3H mouse retina. Science    1974, 186:449-451.-   Farber D B: From mice to men: the cyclic GMP phosphodiesterase gene    in vision and disease. The Proctor Lecture. Invest Ophthalmol Vis    Sci 1995, 36(2):263-275.-   Ferrick D A, Neilson A, Beeson C. Advances in measuring cellular    bioenergetics using extracellular flux. Drug Discov Today. 2008    March; 13(5-6):268-74. doi: 10.1016/j.drudis.2007.12.008. Epub 2008    Feb. 13. Review. PubMed PMID: 18342804.-   Fox D A, Poblenz A T, He L: Calcium overload triggers rod    photoreceptor apoptotic cell death in chemical-induced and inherited    retinal degenerations. Ann NY Acad Sci 1999, 893:282-285.-   Gilliam J C, Chang J T, Sandoval I M, Zhang Y, Li T, Pittler S J,    Chiu W, Wensel T G. Three-dimensional architecture of the rod    sensory cilium and its disruption in retinal neurodegeneration.    Cell. 2012 Nov. 21; 151(5):1029-41. doi: 10.1016/j.cell.2012.10.038.    PubMed PMID: 23178122.-   Graymore C: Metabolism of the Developing Retina. 7. Lactic    Dehydrogenase Isoenzyme in the Normal and Degenerating Retina. a    Preliminary Communication. Exp Eye Res 1964, 89:5-8.-   Hartong D T, Dange M, McGee T L, Berson E L, Dryja T P, Colman R F.    Insights from retinitis pigmentosa into the roles of isocitrate    dehydrogenases in the Krebs cycle. Nat. Genet. 2008 October;    40(10):1230-4. doi: 10.1038/ng.223. Epub 2008 Sep. 21. PubMed PMID:    18806796; PubMed Central PMCID: PMC2596605.-   Ho C H, Piotrowski J, Dixon S J, Baryshnikova A, Costanzo M,    Boone C. Combining functional genomics and chemical biology to    identify targets of bioactive compounds. Curr Opin Chem. Biol. 2011    February; 15(1):66-78. doi: 10.1016/j.cbpa.2010.10.023. Epub 2010    Nov. 17. Review. PubMed PMID: 21093351.-   Ibebunjo C, Chick J M, Kendall T, Eash J K, Li C, Zhang Y, Vickers    C, Wu Z, Clarke B A, Shi J, Cruz J, Fournier B, Brachat S,    Gutzwiller S, Ma Q, Markovits J, Broome M, Steinkrauss M, Skuba E,    Galarneau J R, Gygi S P, Glass D J. Genomic and proteomic profiling    reveals reduced mitochondrial function and disruption of the    neuromuscular junction driving rat sarcopenia. Mol Cell Biol. 2013    January; 33(2):194-212. doi: 10.1128/MCB.01036-12. Epub 2012    Oct. 29. PubMed PMID: 23109432.-   Jaliffa C, Ameqrane I, Dansault A, Leemput J, Vieira V, Lacassagne    E, Provost A, Bigot K, Masson C, Menasche M, Abitbol M. Sirtl    involvement in rd10 mouse retinal degeneration. Invest Ophthalmol    Vis Sci. 2009 August; 50(8):3562-72. doi: 10.1167/iovs.08-2817. Epub    2009 Apr. 30. PubMed PMID: 19407027.-   Jarrett S G, Rohrer B, Perron N R, Beeson C, Boulton M E. Assessment    of mitochondrial damage in retinal cells and tissues using    quantitative polymerase chain reaction for mitochondrial DNA damage    and extracellular flux assay for mitochondrial respiration activity.    Methods Mol Biol. 2013; 935:227-43. doi:    10.1007/978-1-62703-080-9_(—)16. PubMed PMID: 23150372.-   Jewett J C, Bertozzi C R. Cu-free click cycloaddition reactions in    chemical biology. Chem Soc Rev. 2010 April; 39(4):1272-9. Review.    PubMed PMID: 20349533; PubMed Central PMCID: PMC2865253.-   Kanan Y, Moiseyev G, Agarwal N, Ma J X, Al-Ubaidi M R. Light induces    programmed cell death by activating multiple independent proteases    in a cone photoreceptor cell line. Invest Ophthalmol Vis Sci. 2007    January; 48(1):40-51. PubMed PMID: 17197514.-   Kandpal R P, Rajasimha H K, Brooks M J, Nellissery J, Wan J, Qian J,    Kern T S, Swaroop A. Transcriptome analysis using next generation    sequencing reveals molecular signatures of diabetic retinopathy and    efficacy of candidate drugs. Mol Vis. 2012; 18:1123-46. Epub 2012    May 2. PubMed PMID: 22605924; PubMed Central PMCID: PMC3351417.-   Karbowski M, Neutzner A. Neurodegeneration as a consequence of    failed mitochondrial maintenance. Acta Neuropathol. 2012 February;    123 (2): 157-71. doi: 10.1007/s00401-011-0921-0. Epub 2011 Dec. 7.    Review. PubMed PMID: 22143516-   Kroeger H, Messah C, Ahern K, Gee J, Joseph V, Matthes M T, Yasumura    D, Gorbatyuk M S, Chiang W C, Lavail M M, Lin J H. Induction of    Endoplasmic Reticulum Stress Genes, BiP and Chop, in Genetic and    Environmental Models of Retinal Degeneration. Invest Ophthalmol Vis    Sci. 2012 Nov. 9; 53(12):7590-9. doi: 10.1167/iovs.12-10221. PubMed    PMID: 23074209; PubMed Central PMCID: PMC3495601.-   Krysko D V, Agostinis P, Krysko O, Garg A D, Bachert C, Lambrecht B    N, Vandenabeele P. Emerging role of damage-associated molecular    patterns derived from mitochondria in inflammation. Trends Immunol.    2011 April; 32(4):157-64. doi: 10.1016/j.it.2011.01.005. Epub 2011    Feb. 19. Review. PubMed PMID: 21334975.-   Kunchithapautham K, Rohrer B: Apoptosis and Autophagy in    Photoreceptors Exposed to Oxidative Stress. Autophagy 2007, 3(5).-   Lenz E M, Wilson I D: Analytical strategies in metabonomics. J    Proteome Res 2007, 6(2):443-458.-   Lin J H, Lavail M M. Misfolded proteins and retinal dystrophies. Adv    Exp Med. Biol. 2010; 664:115-21. doi:    10.1007/978-1-4419-1399-9_(—)14. Review. PubMed PMID: 20238009;    PubMed Central PMCID: PMC2955894.-   Liu Q, Tan G, Levenkova N, Li T, Pugh E N Jr, Rux J J, Speicher D W,    Pierce E A. The proteome of the mouse photoreceptor sensory cilium    complex. Mol Cell Proteomics. 2007 August; 6(8):1299-317. Epub 2007    May 9. PubMed PMID: 17494944; PubMed Central PMCID: PMC2128741.-   Liu Q, Zhang Q, Pierce E A. Photoreceptor sensory cilia and    inherited retinal degeneration. Adv Exp Med. Biol. 2010; 664:223-32.    doi: 10.1007/978-1-4419-1399-9_(—)26. Review. PubMed PMID: 20238021;    PubMed Central PMCID: PMC2888132.-   Lohr H R, Kuntchithapautham K, Sharma A K, Rohrer B: Multiple,    parallel cellular suicide mechanisms participate in photoreceptor    cell death. Exp Eye Res 2006, 83(2):380-389.-   Lohr H R, Kuntchithapautham K, Sharma A K, Rohrer B. Multiple,    parallel cellular suicide mechanisms participate in photoreceptor    cell death. Exp Eye Res. 2006 August; 83(2):380-9. Epub 2006    Apr. 19. Erratum in: Exp Eye Res. 2006 December; 83(6):1522. PubMed    PMID: 16626700.-   MacMillan K S, Naidoo J, Liang J, Melito L, Williams N S, Morlock L,    Huntington P J, Estill S J, Longgood J, Becker G L, McKnight S L,    Pieper A A, De Brabander J K, Ready J M. Development of    proneurogenic, neuroprotective small molecules. J Am Chem. Soc. 2011    Feb. 9; 133(5):1428-37. doi: 10.1021/ja108211m. Epub 2011 Jan. 6.    PubMed PMID: 21210688; PubMed Central PMCID: PMC3033481.-   Mamidyala S K, Finn M G. In situ click chemistry: probing the    binding landscapes of biological molecules. Chem Soc Rev. 2010    April; 39(4):1252-61. doi: 10.1039/b901969n. Epub 2010 Mar. 1.    Review. PubMed PMID: 20309485.-   Mandal M N, Patlolla J M, Zheng L, Agbaga M P, Tran J T, Wicker L,    Kasus-Jacobi A, Elliott M H, Rao C V, Anderson R E. Curcumin    protects retinal cells from light- and oxidant stress-induced cell    death. Free Radic Biol Med. 2009 Mar. 1; 46(5):672-9. doi:    10.1016/j.freeradbiomed.2008.12.006. Epub 2008 Dec. 24. PubMed PMID:    19121385; PubMed Central PMCID: PMC2810836.-   Marina N, Sajic M, Bull N D, Hyatt A J, Berry D, Smith K J, Martin    K R. Lamotrigine monotherapy does not provide protection against the    loss of optic nerve axons in a rat model of ocular hypertension. Exp    Eye Res. 2012 November; 104:1-6. doi: 10.1016/j.exer.2012.09.002.    Epub 2012 Sep. 13. PubMed PMID: 22982756.-   Mattson M P, Kroemer G: Mitochondria in cell death: novel targets    for neuroprotection and cardioprotection. Trends Mol Med 2003,    9(5):196-205.-   McKnight S L. Back to the future: molecular biology meets    metabolism. Cold Spring Harb Symp Quant Biol. 2011; 76:403-11. doi:    10.1101/sqb.2012.76.013722. Epub 2012 Apr. 17. Review. PubMed PMID:    22510749.-   Mueller E E, Schaier E, Brunner S M, Eder W, Mayr J A, Egger S F,    Nischler C, Oberkofler H, Reitsamer H A, Patsch W, Sperl W,    Kofler B. Mitochondrial haplogroups and control region polymorphisms    in age-related macular degeneration: a case-control study. PLoS One.    2012; 7(2):e30874. doi: 10.1371/journal.pone.0030874. Epub 2012    Feb. 13. PubMed PMID: 22348027; PubMed Central PMCID: PMC3278404.-   Mulkidjanian A Y, Galperin M Y, Makarova K S, Wolf Y I, Koonin E V.    Evolutionary primacy of sodium bioenergetics. Biol Direct. 2008 Apr.    1; 3:13. doi: 10.1186/1745-6150-3-13. PubMed PMID: 18380897; PubMed    Central PMCID: PMC2359735.-   Nicholas P C, Kim D, Crews F T, Macdonald J M: (1)H NMR-Based    Metabolomic Analysis of Liver, Serum, and Brain Following Ethanol    Administration in Rats. Chem. Res Toxicol 2007.-   Nixon E, Simpkins J W. Neuroprotective effects of nonfeminizing    estrogens in retinal photoreceptor neurons. Invest Ophthalmol Vis    Sci. 2012 Jul. 12; 53(8):4739-47. doi: 10.1167/iovs.12-9517. Print    2012 July PubMed PMID: 22700711.-   O'Toole J F, Liu Y, Davis E E, Westlake C J, Attanasio M, Otto E A,    Seelow D, Nurnberg G, Becker C, Nuutinen M, Kärppä M, Ignatius J,    Uusimaa J, Pakanen S, Jaakkola E, van den Heuvel L P, Fehrenbach H,    Wiggins R, Goyal M, Zhou W, Wolf M T, Wise E, Helou J, Allen S J,    Murga-Zamalloa C A, Ashraf S, Chaki M, Heering a S, Chernin G,    Hoskins B E, Chaib H, Gleeson J, Kusakabe T, Suzuki T, Isaac R E,    Quarmby L M, Tennant B, Fujioka H, Tuominen H, Hassinen I, Lohi H,    van Houten J L, Rotig A, Sayer J A, Rolinski B, Freisinger P,    Madhavan S M, Herzer M, Madignier F, Prokisch H, Nurnberg P, Jackson    P K, Khanna H, Katsanis N, Hildebrandt F. Individuals with mutations    in XPNPEP3, which encodes a mitochondrial protein, develop a    nephronophthisis-like nephropathy. J Clin Invest. 2010 March;    120(3):791-802. doi: 10.1172/JCI40076. Epub 2010 Feb. 22. Erratum    in: J Clin Invest. 2010 April; 120(4):1362. Jackson, Peter    [corrected to Jackson, Peter K]. PubMed PMID: 20179356; PubMed    Central PMCID: PMC2827951.-   Osborne N N, Del Olmo-Aguado S. Maintenance of retinal ganglion cell    mitochondrial functions as a neuroprotective strategy in glaucoma.    Curr Opin Pharmacol. 2012 Sep. 19. doi:pii: 51471-4892(12)00159-2.    10.1016/j.coph.2012.09.002. [Epub ahead of print] PubMed PMID:    22999653.-   Pappas D J, Gabatto P A, Oksenberg D, Khankhanian P, Baranzini S E,    Gan L, Oksenberg J R. Transcriptional expression patterns triggered    by chemically distinct neuroprotective molecules. Neuroscience. 2012    Dec. 13; 226:10-20. doi: 10.1016/j.neuroscience.2012.09.007. Epub    2012 Sep. 15. PubMed PMID: 22986168; PubMed Central PMCID:    PMC3489981.-   Pereira D A, Williams J A. Origin and evolution of high throughput    screening. Br J. Pharmacol. 2007 September; 152(1):53-61. Epub 2007    Jul. 2. Review. PubMed PMID: 17603542; PubMed Central PMCID:    PMC1978279.-   Perron N R, Beeson C, Rohrer B. Early alterations in mitochondrial    reserve capacity; a means to predict subsequent photoreceptor cell    death. J Bioenerg Biomembr. 2012 Oct. 23. [Epub ahead of print]    PubMed PMID: 23090843.-   Pieper A A, Xie S, Capota E, Estill S J, Zhong J, Long J M, Becker G    L, Huntington P, Goldman S E, Shen C H, Capota M, Britt J K, Kotti    T, Ure K, Brat D J, Williams N S, MacMillan K S, Naidoo J, Melito L,    Hsieh J, De Brabander J, Ready J M, McKnight S L. Discovery of a    proneurogenic, neuroprotective chemical. Cell. 2010 Jul. 9;    142(1):39-51. doi: 10.1016/j.cell.2010.06.018. PubMed PMID:    20603013; PubMed Central PMCID: PMC2930815.-   Pierce E A, Quinn T, Meehan T, McGee T L, Berson E L, Dryja T P:    Mutations in a gene encoding a new oxygen-regulated photoreceptor    protein cause dominant retinitis pigmentosa. Nat Genet. 1999,    22(3):248-254.-   Pierce E A: Pathways to photoreceptor cell death in inherited    retinal degenerations. Bioessays 2001, 23(7):605-618.-   Qin L X, Beyer R P, Hudson F N, Linford N J, Morris D E, Kerr K F.    Evaluation of methods for oligonucleotide array data via    quantitative real-time PCR. BMC Bioinformatics. 2006 Jan. 17; 7:23.    PubMed PMID: 16417622; PubMed Central PMCID: PMC1360686.-   Rezaie T, McKercher S R, Kosaka K, Seki M, Wheeler L, Viswanath V,    Chun T, Joshi R, Valencia M, Sasaki S, Tozawa T, Satoh T, Lipton    S A. Protective effect of carnosic Acid, a pro-electrophilic    compound, in models of oxidative stress and light-induced retinal    degeneration. Invest Ophthalmol Vis Sci. 2012 Nov. 27;    53(12):7847-54. doi: 10.1167/iovs.12-10793. PubMed PMID: 23081978;    PubMed Central PMCID: PMC3508754.-   Richards A J, Muller B, Shotwell M, Cowart L A, Rohrer B, Lu X.    Assessing the functional coherence of gene sets with metrics based    on the Gene Ontology graph. Bioinformatics. 2010 Jun. 15;    26(12):179-87. doi: 10.1093/bioinformatics/btq203. PubMed PMID:    20529941; PubMed Central PMCID: PMC2881388.-   Richards T A, Cavalier-Smith T. Myosin domain evolution and the    primary divergence of eukaryotes. Nature. 2005 Aug. 25;    436(7054):1113-8. PubMed PMID: 16121172.-   Rohrer B, Matthes M T, LaVail M M, Reichardt L F: Lack of p75    receptor does not protect photoreceptors from light-induced cell    death. Exp Eye Res 2003, 76(1):125-129-   Rohrer B, Pinto F R, Hulse K E, Lohr H R, Zhang L, Almeida J S.    Multidestructive pathways triggered in photoreceptor cell death of    the rd mouse as determined through gene expression profiling. J    Biol. Chem. 2004 Oct. 1; 279(40):41903-10. Epub 2004 Jun. 24. PubMed    PMID: 15218024.-   Ronquillo C C, Bernstein P S, Baehr W. Senior-Løken syndrome: A    syndromic form of retinal dystrophy associated with    nephronophthisis. Vision Res. 2012 Dec. 15; 75:88-97. doi:    10.1016/j.visres.2012.07.003. Epub 2012 Jul. 20. PubMed PMID:    22819833; PubMed Central PMCID: PMC3504181.-   Sancho-Pelluz J, Alavi M V, Sahaboglu A, Kustermann S, Farinelli P,    Azadi S, van Veen T, Romero F J, Paquet-Durand F, Ekstrom P.    Excessive HDAC activation is critical for neurodegeneration in the    rd1 mouse. Cell Death Dis. 2010; 1:e24. doi: 10.1038/cddis.2010.4.    PubMed PMID: 21364632; PubMed Central PMCID: PMC3032332.-   Sancho-Pelluz J, Arango-Gonzalez B, Kustermann S, Romero F J, van    Veen T, Zrenner E, Ekström P, Paquet-Durand F. Photoreceptor cell    death mechanisms in inherited retinal degeneration. Mol Neurobiol.    2008 December; 38(3):253-69. doi: 10.1007/s12035-008-8045-9. Epub    2008 Nov. 4. Review. PubMed PMID: 18982459.-   SanGiovanni J P, Arking D E, Iyengar S K, Elashoff M, Clemons T E,    Reed G F, Henning A K, Sivakumaran T A, Xu X, DeWan A, Agron E,    Rochtchina E, Sue C M, Wang J J, Mitchell P, Hoh J, Francis P J,    Klein M L, Chew E Y, Chakravarti A. Mitochondrial DNA variants of    respiratory complex I that uniquely characterize haplogroup T2 are    associated with increased risk of age-related macular degeneration.    PLoS One. 2009; 4(5):e5508. doi: 10.1371/journal.pone.0005508. Epub    2009 May 12. PubMed PMID: 19434233; PubMed Central PMCID:    PMC2677106.-   Schrier S A, Falk M J. Mitochondrial disorders and the eye. Curr    Opin Ophthalmol. 2011 September; 22(5):325-31. doi:    10.1097/ICU.0b013e328349419d. Review. PubMed PMID: 21730846.-   Sharma A K, Rohrer B: Calcium-induced calpain mediates apoptosis via    caspase-3 in a mouse photoreceptor cell line. J Biol Chem 2004,    279(34):35564-35572.-   Sharma A K, Rohrer B. Calcium-induced calpain mediates apoptosis via    caspase-3 in a mouse photoreceptor cell line. J Biol. Chem. 2004    Aug. 20; 279(34):35564-72. Epub 2004 Jun. 18. PubMed PMID: 15208318.-   Sharma A K, Rohrer B. Sustained elevation of intracellular cGMP    causes oxidative stress triggering calpain-mediated apoptosis in    photoreceptor degeneration. Curr Eye Res. 2007 March; 32(3):259-69.    PubMed PMID: 17453946.-   Shimazaki H, Hironaka K, Fujisawa T, Tsuruma K, Tozuka Y, Shimazawa    M, Takeuchi H, Hara H. Edaravone-loaded liposome eyedrops protect    against light-induced retinal damage in mice. Invest Ophthalmol Vis    Sci. 2011 Sep. 21; 52(10):7289-97. doi: 10.1167/iovs.11-7983. Print    2011 Sep. PubMed PMID: 21849425.-   Smith J J, Kenney R D, Gagne D J, Frushour B P, Ladd W, Galonek H L,    Israelian K, Song J, Razvadauskaite G, Lynch A V, Carney D P,    Johnson R J, Lavu S, Iffland A, Elliott P J, Lambert P D, Elliston K    O, Jirousek M R, Milne J C, Boss O. Small molecule activators of    SIRT1 replicate signaling pathways triggered by calorie restriction    in vivo. BMC Syst Biol. 2009 Mar. 10; 3:31. doi:    10.1186/1752-0509-3-31. PubMed PMID: 19284563; PubMed Central PMCID:    PMC2660283.-   Spinazzi M, Cazzola S, Bortolozzi M, Baracca A, Loro E, Casarin A,    Solaini G, Sgarbi G, Casalena G, Cenacchi G, Malena A, Frezza C,    Carrara F, Angelini C, Scorrano L, Salviati L, Vergani L. A novel    deletion in the GTPase domain of OPA1 causes defects in    mitochondrial morphology and distribution, but not in function. Hum    Mol Genet. 2008 Nov. 1; 17(21):3291-302. doi: 10.1093/hmg/ddn225.    Epub 2008 Aug. 4. PubMed PMID: 18678599.-   Stone J, Maslim J, Valter-Kocsi K, Mervin K, Bowers F, Chu Y,    Barnett N, Provis J, Lewis G, Fisher S K et al: Mechanisms of    photoreceptor death and survival in mammalian retina. Prog Retin Eye    Res 1999, 18(6):689-735.-   Sumantran V N. Cellular chemosensitivity assays: an overview.    Methods Mol Biol. 2011; 731:219-36. doi:    10.1007/978-1-61779-080-5_(—)19. Review. PubMed PMID: 21516411.-   Tan E, Ding X Q, Saadi A, Agarwal N, Naash M I, Al-Ubaidi M R:    Expression of cone-photoreceptor-specific antigens in a cell line    derived from retinal tumors in transgenic mice. Invest Ophthalmol    Vis Sci 2004, 45(3):764-768.-   Tan E, Ding X Q, Saadi A, Agarwal N, Naash M I, Al-Ubaidi M R.    Expression of cone-photoreceptor-specific antigens in a cell line    derived from retinal tumors in transgenic mice. Invest Ophthalmol    Vis Sci. 2004 March; 45(3):764-8. PubMed PMID: 14985288; PubMed    Central PMCID: PMC2937568.-   Tesla R, Wolf H P, Xu P, Drawbridge J, Estill S J, Huntington P,    McDaniel L, Knobbe W, Burket A, Tran S, Starwalt R, Morlock L,    Naidoo J, Williams N S, Ready J M, McKnight S L, Pieper A A.    Neuroprotective efficacy of aminopropyl carbazoles in a mouse model    of amyotrophic lateral sclerosis. Proc Natl Acad Sci USA. 2012 Oct.    16; 109(42):17016-21. doi: 10.1073/pnas.1213960109. Epub 2012    Oct. 1. PubMed PMID: 23027932; PubMed Central PMCID: PMC3479516.-   Travis G H: Mechanisms of cell death in the inherited retinal    degenerations. Am J Hum Genet. 1998, 62(3):503-508.-   Trifunović D, Sahaboglu A, Kaur J, Mend S, Zrenner E, Ueffing M,    Arango-Gonzalez B, Paquet-Durand F. Neuroprotective strategies for    the treatment of inherited photoreceptor degeneration. Curr Mol Med.    2012 June; 12(5):598-612. Review. PubMed PMID: 22515977.-   Tu B P, Mohler R E, Liu J C, Dombek K M, Young E T, Synovec R E,    McKnight S L. Cyclic changes in metabolic state during the life of a    yeast cell. Proc Natl Acad Sci USA. 2007 Oct. 23; 104(43):16886-91.    Epub 2007 Oct. 16. PubMed PMID: 17940006; PubMed Central PMCID:    PMC2040445.-   Van Bergen N J, Crowston J G, Kearns L S, Staffieri S E, Hewitt A W,    Cohn A C, Mackey D A, Trounce I A. Mitochondrial oxidative    phosphorylation compensation may preserve vision in patients with    OPA1-linked autosomal dominant optic atrophy. PLoS One. 2011;    6(6):e21347. doi: 10.1371/journal.pone.0021347. Epub 2011 Jun. 22.    PubMed PMID: 21731710; PubMed Central PMCID: PMC3120866.-   Vingolo E M, De Mattia G, Giusti C, Forte R, Laurenti O, Pannarale M    R: Treatment of nonproliferative diabetic retinopathy with    Defibrotide in noninsulin-dependent diabetes mellitus: a pilot    study. Acta Ophthalmol Scand 1999, 77(3):315-320.-   Wenzel A, Grimm C, Samardzija M, Reme C E: Molecular mechanisms of    light-induced photoreceptor apoptosis and neuroprotection for    retinal degeneration. Prog Retin Eye Res 2005, 24(2):275-306.-   Whitfield J F, Chakravarthy B R. The neuronal primary cilium: driver    of neurogenesis and memory formation in the hippocampal dentate    gyrus Cell Signal. 2009 September; 21(9):1351-5. doi:    10.1016/j.cellsig.2009.02.013. Epub 2009 Feb. 26. Review. PubMed    PMID: 19249355.-   Winkler B S, Pourcho R G, Starnes C, Slocum J, Slocum N. Metabolic    mapping in mammalian retina: a biochemical and 3H-2-deoxyglucose    autoradiographic study. Exp Eye Res. 2003 September; 77(3):327-37.    PubMed PMID: 12907165.-   Winkler B S. Letter to the editor: Comments on retinal metabolic    state in P23H and normal retinas. Am J Physiol Cell Physiol. 2010    July; 299(1):C185; author reply C186-7. doi:    10.1152/ajpcell.00109.2010. PubMed PMID: 20554913.-   Yamada Y, Hidefumi K, Shion H, Oshikata M, Haramaki Y. Distribution    of chloroquine in ocular tissue of pigmented rat using    matrix-assisted laser desorption/ionization imaging quadrupole    time-of-flight tandem mass spectrometry. Rapid Commun Mass SpectroM.    2011 Jun. 15; 25(11):1600-8. doi: 10.1002/rcm.5021. PubMed PMID:    21594935.-   Yang L, Nyalwidhe J O, Guo S, Drake R R, Semmes O J. Targeted    identification of metastasis-associated cell-surface    sialoglycoproteins in prostate cancer. Mol Cell Proteomics. 2011    June; 10(6):M110.007294. doi: 10.1074/mcp.M110.007294. Epub 2011    Mar. 29. PubMed PMID: 21447706; PubMed Central PMCID: PMC3108840.-   Ying W. NAD+ and NADH in cellular functions and cell death. Front    Biosci. 2006 Sep. 1; 11:3129-48. Review. PubMed PMID: 16720381.-   Farber, D. B., From mice to men: the cyclic GMP phosphodiesterase    gene in vision and disease. The Proctor Lecture. Invest. Ophthalmol.    Vis. Sci., 1995. 36(2): p. 263-275.-   Farber, D. B. and R. N. Lolley, Cyclic guanosine monophosphate:    elevation in degenerating photoreceptor cells of the C3H mouse    retina. Science, 1974. 186: p. 449-451.-   Fox, D. A., A. T. Poblenz, and L. He, Calcium overload triggers rod    photoreceptor apoptotic cell death in chemical-induced and inherited    retinal degenerations. Ann. N.Y. Acad. Sci., 1999. 893: p. 282-285.-   Ogilvie, J. M., et al., A reliable method for organ culture of    neonatal mouse retina with long-term survival. J. Neurosci.    Methods, 1999. 87(1): p. 57-65.

It is to be understood that the invention is not limited to theparticular embodiments of the invention described above, as variationsof the particular embodiments may be made and still fall within thescope of the appended claims.

What is claimed is:
 1. The compound according to formula (I):

wherein: R₁ is pyridinyl; one of R₂ or R_(2′) is hydrogen and the otheris —CH₂R₃ or —CH₂C(O)R₃; and R₃ unsubstituted phenyl or phenyl mono-,bi- or tri-substituted independently with alkoxy, or a pharmaceuticallyacceptable salt thereof.
 2. The compound according to claim 1, whereinone of R₂ or R_(2′) is hydrogen and the other is —CH₂R₃.
 3. The compoundaccording to claim 1, wherein one of R₂ or R_(2′) is hydrogen and theother is —CH₂C(O)R₃.
 4. The compound according to claim 1, wherein R₃ isphenyl mono-substituted with methoxy.
 5. The compound according to claim1, wherein R₃ is phenyl bi-substituted with methoxy.
 6. The compoundaccording to claim 1, wherein R₃ is phenyl tri-substituted with methoxy.7. The compound according to claim 1, wherein R₂ is hydrogen, R_(2′) is—CH₂C(O)R₃ and R₃ is phenyl bi-substituted with methoxy.
 8. The compoundaccording to claim 1, wherein said compound is: Ethyl1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate;Ethyl1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-5-(pyridin-2-yl)-1H-pyrazole-3-carboxylate;Ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate;Ethyl1-(2-(2,4-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-3-yl)-1H-pyrazole-5-carboxylate;or Ethyl1-(2-(2,5-dimethoxyphenyl)-2-oxoethyl)-3-(pyridin-2-yl)-1H-pyrazole-5-carboxylate.9. A pharmaceutical composition, comprising a therapeutically effectiveamount of a compound according to claim 1 or a pharmaceuticallyacceptable salt thereof and a pharmaceutically acceptable carrier.